MicroRNA-711-Induced Downregulation of Angiopoietin-1 Mediates Neuronal Cell Death.

Abstract

Angiopoietin-1 (Ang-1) is a well-known endothelial growth factor, but its effects on neurons have yet to be elucidated. We show that Ang-1 is rapidly downregulated in the injured brain after controlled cortical impact (CCI), a mouse experimental traumatic brain injury (TBI) model and in etoposide-induced neuronal apoptosis in vitro. Ang-1 treatment inhibits etoposide-induced upregulation of proapoptotic B-cell lymphoma 2 (Bcl-2) family members Noxa, p53 upregulated modulator of apoptosis (Puma), Bcl-2 interacting mediator of cell death (Bim), and Bcl-2-associated X protein (Bax); reduces markers of caspase-dependent (cytochrome c release/caspase activation) and caspase-independent (apoptosis-inducing factor release) pathways; and limits neuronal cell death. Ang-1 treatment phosphorylates receptors Tunica interna endothelial cell kinase 2 (Tie2), and β1-integrin and limits the etoposide-induced decrease in protein kinase B (Akt) activity. Blocking Tie2 and β1-integrin signaling reduces Ang-1 neuroprotective effects. After both TBI and etoposide treatment microRNA (miR)-711 are upregulated, consistent with its putative role as a negative regulator of Ang-1. We show that miR-711 directly targets the Ang-1 messenger RNA (mRNA), decreasing Ang-1 expression. Increased levels of miR-711 and Ang-1 mRNA are found in the RNA-induced silencing complex complex site of miR-mediated degradation of target mRNAs after etoposide treatment and the miR-711mimic downregulates Ang-1. Administration of miR-711 inhibitor elevates Ang-1 after TBI whereas Ang-1 administration increases Akt activation; reduces Puma, Noxa, Bim, and Bax levels; and attenuates caspase-dependent and -independent neuronal apoptosis 24 h after TBI. Ang-1 also attenuates neuronal degeneration, increases gene expression of molecules that maintain blood-brain barrier integrity, and reduces post-traumatic lesion volume/edema 24 h after TBI. Although we only observed short-term neuroprotective effects after Ang-1 administration, miR-711-dependent downregulation of Ang-1, followed by Akt pathway inhibition, may play a role in neuronal cell death after neuronal injury in vitro and after experimental TBI.